Polymorphic form of reduced beta-nicotinamide mononucleotide disodium salt, and preparation method therefor and use thereof

The present invention relates to the field of chemical raw materials for pharmaceuticals, healthcare product and cosmetics, and specifically relates to the polymorphic forms of reduced β-nicotinamide mononucleotide disodium salts, and preparation method therefor and use thereof.

Nicotinamide adenine dinucleotide (NAD), one of the hottest molecules in the field of anti-aging, is without exception at the center of successive generations of anti-aging substances. NADis an essential coenzyme required for over 500 enzymatic reactions and is well known for its role in oxidation and reduction (Ansari and Raghava, 2010; Rajman et al. 2018; Stein and Imai, 2012). A growing number of studies have instructed that increasing NADequivalents significantly improves multiple organ functions, including liver function, kidney function, heart function and skeletal muscle function (Canto et al. 2012; Mills et al. 2016; Rajman et al., 2018). NADcan be synthesized using tryptophan in the de novo biosynthesis pathway (de novo biosynthesis pathway), nicotinic acid (NA) in the preiss-handler pathway, as well as nicotinamide (NAM), nicotinamide riboside (NR), and nicotinamide mononucleotide (NMN) in the salvage pathway (Canto et al. 2015; Chiarugi et al. 2012; Johnson and Imai 2018). In particular, as key intermediates of NAD, NAM, NR, and NMN have been extensively studied for their potential therapeutic roles in many mouse disease models (Mills et al., 2016), wherein NMN is considered to be the most suitable NADprecursor at present, and NMN is currently hotly sold in the global marketplace and is highly favored by consumers.

NMNH (molecular structure as shown in formula (A)), known as “reduced nicotinamide mononucleotide” or “reduced β-nicotinamide mononucleotide” in Chinese, is a reduced form of NMN. NMNH is a new precursor for NADsupplementation with better NAD-promoting effects than NMN, as well as other biological functions such as increasing cellular antioxidant capacity, reducing fat accumulation, reducing inflammatory response and inhibiting tumor cell growth, etc., and is a health-promoting reagent with significant commercial potential (WO2021098725A1).

The synthesis process of NMNH in the market is still in the laboratory research and development stage, and it is not yet possible to realize industrial production. The main technical difficulties in the synthesis of NMNH are: 1) NMNH is a reduced form of NMN, which is easily oxidized by air; 2) In the laboratory, an aqueous solution with higher purity can be obtained only by preparative chromatography, and an amorphous solid can be obtained by freeze-drying. This method has no purification effect on the product, and the amorphous solid obtained by freeze-drying is foamy, has poor fluidity, is easy to absorb moisture and becomes oil, and degrades quickly; 3) No polymorphic form has been reported. It is well known that freeze-drying is not used industrially unless it is necessary because of its high energy consumption and limited production capacity; amorphous solids have higher energy and are more unstable than crystalline solids.

Therefore, there is an urgent need in this field to develop new NMNH compounds and polymorphic form thereof with advantages of good stability, good fluidity and suitability for industrialization.

The purpose of the present invention is to provide new NMNH compounds and polymorphic forms thereof, namely polymorphic forms of NMNH disodium salts and preparation method therefor and the uses thereof, which have the advantages of good stability, good fluidity and suitability for industrialization.

A first aspect of the present invention provides crystals of a reduced β-nicotinamide mononucleotide disodium salt, and the crystal form of the crystals is selected from the group consisting of: crystal form A, crystal form B, and crystal form C.

In another preferred embodiment, the structure of the reduced β-nicotinamide mononucleotide disodium salt is shown in Formula I:

In another preferred embodiment, the crystals of reduced β-nicotinamide mononucleotide disodium salt are hydrate.

In another preferred embodiment, the structure of the reduced β-nicotinamide mononucleotide disodium salt is shown in Formula II

In another preferred embodiment, n is an integer or a non-integer.

In another preferred embodiment, n is a positive integer of ≥2, preferably 2-10, more preferably 5-9.

In another preferred embodiment, the XRPD pattern of the crystal form A comprises 3 or more 2θ values selected from the group consisting of: 12.7°±0.2°, 15.9°±0.2°, 18.0°±0.2°, 20.4°±0.2°, 20.9°±0.2°, and 31.8°±0.2°.

In another preferred embodiment, the XRPD pattern of the crystal form A comprises 1 or more 2θ values selected from the group consisting of: 10.5°±0.2°, 19.8°±0.2°, 22.6°±0.2°, 24.0°±0.2°, 26.1°±0.2°, 28.7°±0.2°, 30.8°±0.2°, and 33.4°±0.2°.

In another preferred embodiment, the crystal form A further has one or more features selected from the following group:

In another preferred embodiment, the crystal form B has one or more features selected from the following group:

In another preferred embodiment, the crystal form C has one or more features selected from the following group:

A second aspect of the present invention provides a method for preparing crystals of reduced β-nicotinamide mononucleotide disodium salt, the method comprises the following steps:

In another preferred embodiment, the first solvent and the second solvent are the same or different, and are independently selected from the group consisting of: water, acetonitrile, tetrahydrofuran, methyl tert-butyl ether, 2-methyltetrahydrofuran, dichloromethane, N,N-dimethylformamide, N,N-dimethylacetamide, N,N-diethylformamide, dimethylsulfoxide, ethyl acetate, isopropyl acetate, ketone solvent, alcohol solvent, or a combination thereof.

In another preferred embodiment, the ketone solvent is selected from the group consisting of: acetone, 2-butanone, methyl isobutyl ketone, methyl tert-butyl ketone, 3-methyl-2-butanone, or a combination thereof.

In another preferred embodiment, the alcohol solvent is selected from the group consisting of: methanol, ethanol, n-propanol, isopropanol, n-butanol, isobutanol, n-amyl alcohol.

In another preferred embodiment, the method further comprises: step 3), drying the crystals.

In another preferred embodiment, the drying comprises: drying the resulting crystals under vacuum for 2-30 hours

In another preferred embodiment, the crystal is crystal form A.

In another preferred embodiment, when the crystals obtained in step 2) are crystal form B and/or crystal form C, then the step 2) further comprises a sub-step 2a), wherein the crystal form B and/or the crystal form C are placed in a moisture-containing gas and thus converted to crystal form A.

In another preferred embodiment, in step 2a), crystal form B and/or crystal form C are placed in air and thus converted to crystal form A.

A third aspect of the present invention provides a composition, the composition comprises: (a) any of the crystals described in the first aspect, and (b) pharmaceutically acceptable excipients or carriers, acceptable excipients or carriers for healthcare product, acceptable excipients or carriers for cosmetic, or acceptable excipients or carriers for food.

In another preferred embodiment, the composition is selected from the group consisting of: pharmaceutical compositions, healthcare product compositions, cosmetic compositions, or food compositions.

In another preferred embodiment, the pharmaceutical composition comprises (a) any of the crystals described in the first aspect, and (b) pharmaceutically acceptable excipients or carriers.

In another preferred embodiment, the dosage form of the pharmaceutical composition is selected from the group consisting of: oral formulation, injectable dosage form, respiratory delivery dosage form, dermal delivery dosage form, mucosal delivery dosage form, luminal delivery dosage form and the like.

In another preferred embodiment, the healthcare product composition comprises (a) any of the crystals described in the first aspect, and (b) acceptable excipients or carriers for healthcare product.

In another preferred embodiment, the cosmetic composition comprises (a) any of the crystals described in the first aspect, and (b) acceptable excipients or carriers for cosmetic.

In another preferred embodiment, the cosmetic composition comprises a cosmetic of use selected from the group consisting of: skin cosmetic, hair cosmetic, beauty cosmetic, special function cosmetic.

In another preferred embodiment, the food composition comprises (a) any of the crystals described in the first aspect, and (b) acceptable excipients or carriers for food.

A fourth aspect of the present invention provides the use of a crystal in the preparation of a drug or a healthcare product or a cosmetic or a food additive.

In another preferred embodiment, the drug is used for protecting the optic nerve, improving retinal damage, preventing/treating hair loss, preventing/improving cardiovascular and cerebrovascular diseases, inhibiting renal tubular damage and aging, preventing hepatic fibrosis, improving fatty liver disease, improving symptoms of dry eye, repairing renal damage, preventing diabetes/nephropathy, improving symptoms of sarcopenia in the elderly, treating chronic inflammation, alleviating condition of polycystic ovary syndrome patients, preventing/delaying glaucoma, reducing neuroinflammation, reducing cardiotoxicity of anthracycline chemotherapy drugs, aiding recovery from cerebral infarction, and combating heart failure in the elderly.

In another preferred embodiment, the healthcare product are used for slowing down cellular aging, slowing down reproductive aging in females, improving fertility, improving menopause, enhancing male sexual function, improving sleep, soothing mood, boosting energy, improving cardiovascular function, boosting cardiovascular health, boosting immunity, improving sub-health, preventing tumors, preventing Alzheimer's disease, and the like.

In another preferred embodiment, the cosmetic product is used for improving damaged cell function, improving skin/hair texture, preventing/treating photoaging of the skin, maintaining softness and elasticity of the skin, slowing down aging of the skin, and the like.

In another preferred embodiment, the food additive is used for improving appetite, improving digestion, promoting metabolism, promoting hair/nail growth, etc., and enhancing nutritional value.

It should be understood that, within the scope of the present invention, each of the above-described technical features of the present invention and each of the technical features specifically described below (e.g., in the embodiments) can be combined with each other, thereby constituting a new or preferred technical solution. For the sake of limitation of space, we will not repeat all of them herein.

Through extensive and intensive research, the present inventor has for the first time unexpectedly developed a specific salt of reduced NMNH, and the salt is the disodium salt of NMNH. The studies of the present invention have shown that the polymorphic form (particularly crystal form A) of the disodium salt of NMNH, has excellent stability. Compared to its amorphous solid, the amorphous form changes from a solid to an oil or viscous mass and degrades rapidly upon placement. In addition, the polymorphic form of the present invention has the advantages of high purity, good stability, good fluidity, and low hygroscopicity, which makes it suitable for use in pharmaceutical compositions, healthcare products, cosmetics, food additives, and the like. On this basis, the inventor has completed the present invention.

Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the present invention belongs.

As used herein, n in the term “nHO” refers to all possible values including integers and non-integers between 2 and 10; “HO” is the chemical formula for water, representing water molecules or water.

As used herein, when used in reference to a specifically enumerated value, the term “about” means that the value can vary by no more than 1% from the specifically enumerated value, e.g., as used herein, the expression “about 100” includes all values between 99 and 101 (e.g., 99.1, 99.2, 99.3, 99.4, etc.).

As used herein, the terms “contains” or “includes (comprises)” can be open-ended, semi-enclosed and closed-ended. In other words, the terms mentioned also include “substantially consisting of” or “consisting of”.

As used herein, the term “n or more 2θ values selected from the group consisting of” means including n and any positive integer greater than n (e.g., n, n+1, . . . ), where the upper limit Nup is the number of all 20 peaks in the group. For example, “3 or more” includes not only 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, . . . each positive integer of the upper Nup, but also “4 or more”. or more”, “5 or more”, “6 or more” and other ranges.

NMNH Disodium Salt